A variety of in vitro and clinical studies have shown that cancer cells can exhibit resistance to chemotherapeutic drugs, including two drugs commonly used for chemotherapeutic treatment of breast cancer, particularly, paclitaxel and doxorubicin. Moreover, long-term exposure of tumor cells to one chemotherapeutic agent can often result in cross-resistance to a variety of structurally unrelated drugs. This phenomenon is termed multidrug resistance (MDR) [1, 2]. The development of multidrug resistance in tumor cells is believed to be a major obstacle to the treatment of cancer by chemotherapy. One of the most well characterized mechanisms for multidrug resistance in tumor cells involves the increased expression of a superfamily of ATP-binding cassette (ABC) drug transporters. These include such well known proteins as P-glycoprotein (P-gp or ABCB1), the multidrug resistance proteins MRP1 (ABCC1) and MRP2 (ABCC2), and the breast cancer resistance protein (BCRP or ABCG2) [3-9]. These ABC transporters actively transport a variety of structurally unrelated chemotherapeutic drugs out of the cells or into vesicles, thereby decreasing intracellular drug accumulation and inhibiting drug-induced cytotoxicity [10, 11]. Multidrug resistance can also stem from a variety of additional mechanisms [12, 13] including enhanced expression of glutathione-S-transferase or glutathione peroxidase [14], reduced topoisomerase II expression [15, 16], cell adhesion to extracellular matrix proteins [17] and the inhibition of drug-induced apoptosis [18-20].
Attempts have been made to overcome multidrug resistance by inhibiting the expression of genes involved in this phenomenon [21], or by blocking P-gp function using cyclosporin A [22], valspodar (PSC-833) [23-26], or verapamil [27]. These P-gp inhibitors increase cellular drug accumulation in drug-resistant cells by competing for drug-binding sites on P-gp, allowing less drug to be extruded by the transporter [28-30]. P-gp inhibitors may improve the outcome of chemotherapy for some cancers by enabling drugs to remain in tumor cells where they can exert their cytotoxic effects. However, this approach appears to be drug and/or cell type specific. For example, cyclosporin A was able to inhibit multidrug resistance by disturbing P-gp function in doxorubicin-resistant human myeloid leukemia cells [22], daunorubicin-resistant human T-lymphoblastoid cells [1] and vincristine-resistant K562/MDR leukemia cells [31], while having no effect on the cytotoxicity of 6-mercaptopurine or mitomycin C in HeLa or Hvr100-6 cells selected for resistance to these agents [32]. Similarly, valspodar can completely restore the cytotoxicity and intracellular accumulation of paclitaxel in doxorubicin-resistant NCI-ADR cells while having no effect on 5-fluorouracil cytotoxicity and uptake [33]. In addition, valspodar could only partially restore doxorubicin cytotoxicity in doxorubicin-resistant MCF-7 cells, while fully restoring paclitaxel cytotoxicity in paclitaxel-resistant MCF-7 cells.
Thus, there remains a need to identify other agents that have the potential to kill a variety of drug-resistant tumor cells, particularly breast cancer cells. The mechanism of action of these agents would preferably be in a P-glycoprotein-independent manner. An agent which could kill breast tumor cells resistant to the anthracycline doxorubicin or the taxane paclitaxel (even in P-gp-expressing cells) would be highly desirable, considering that these drugs are widely used in the treatment of breast cancer and approximately half of patients respond to taxanes after anthracycline chemotherapy [34].
Calphostin C, a highly specific photoactivatable inhibitor of phorbol-responsive protein kinase C (PKC) isoforms [35], is one such agent which has shown some promise in killing drug resistant tumor cells including daunorubicin-resistant tumor cells [36]. Calphostin C has also been employed as an agent to treat non-drug resistant MCF-7 breast cancer cells [77]. Cell death induced by calphostin C has been shown to be independent of the p53, pRb and p16 status of the cells, suggesting that the mechanism of cell death caused by this reagent is likely unaffected by common genetic alterations in cancer [37, 38]. However, it was not previously known whether calphostin C would be useful as chemotherapeutic agent for multi-dug resistant tumor cells, in particular breast cancer tumors resistant to taxane or anthracycline drugs.